Isolation and identification of phagocytic activity methods for Kupffer cells from neonatal mouse

Objective To establish a convenient method of isolating Kupffer cells from neonatal mouse liver cells and to provide experimental cells for investigating the pathogenesis of Biliary Atresia.Methods A parenchymal hepatocyte enriched fraction was prepared from 6-8 neonatal mouse liver cells and seeded into culture flasks.After 3 to 4 days of culture,when most hepatocytes were vigorously proliferated on the cell sheet.By cell craper,the adherence cells were readily detached and transferred in another plastic dishes.After 45 minutes incubation,the quick and selective adhesion cells were what we wanted:second adherence cells.Incubated with light microscopy (2 μm) for l hour,these second adherence cells were observed under the laser confocal microscope.Results These second adherence cells were identified by their immunoreactivity to the F4/80 antibody,and to 99％ purity.As more than 107 F4/80 positive cells could be recovered simply from the method.The cells possessed active phagocytosis of typical macrophage as light microscopy (2 μm) could be seen in the cytoplasm.Conclusion Our procedure might implicate a novel alternative by selective adhesion to obtain Kupffer cells from neonatal mouse liver cells in sufficient number and purity without complex equipment and skills,which is especially usefully for studying the pathogenesis of Biliary Atresia. Key words: Kupffer cell;  Isolated culture ;  Phagocytosis ;  Neonatal mouse